1. Field of the Invention
This invention relates generally to plasma containing compositions and methods for sterilizing and preserving plasma for storage and transportation at ambient temperatures. The plasma compositions are designed specifically for the trauma care field.
2. Description of the Background Fresh frozen plasma (FFP) is essential for the clinical management of coagulopathies associated with combat trauma. However, a frozen formulation has three major limitations: (a) FFP must be kept frozen at about minus 18° C. or below; (b) the number of FFP units transshipped is restricted by the dry ice (CO2) volume and/or weight limitations of transport; (c) thawing time of intact plasma units is generally 30 to 40 minutes.
Clearly, the limitations of the frozen formulation reduce plasma availability in the field, or in rural locations, for both logistic and practical laboratory reasons. A freeze-dried formulation, alternatively, could remedy the storage and shipment problem by converting liquid plasma into a lightweight, solid product that is stable at ambient temperature. Currently, the advantage of using freeze-dried plasma verses frozen plasma has been mostly recognized in developing countries [1] and [2]. There are limitations in facilities for preparation, transportation and storage of frozen blood products in most of these countries. Therefore, freeze-dried products with less storage and transportation requirements are preferred.
Pooled plasma was lyophilized for the first time during World War II. However, it was discovered that the process of freeze-drying does not kill viruses in plasma. In addition, the use of plasma from large pools carried an unacceptable risk of transmitting pathogens [3]. Consequently, the production of freeze-dried plasma was abandoned.
Several methods for pathogen inactivation in plasma are now at different stages of development. Such methods are based on: solvent/detergent treatment [4]; utilization of vitamin B2, riboflavin and light [5] application of psoralens and UV light [6]. The current endeavor is to freeze-dry pathogen-inactivated plasma products. These products will guarantee both unconstrained plasma availability and safety. Several groups report stability results for freeze-dried, pathogen-inactivated, solvent/detergent (SD)-treated plasma products. Hellstern et al. describe the production of freeze-dried and deep-frozen batches of SD plasma, and characterize the product in vitro [7]. Clotting factors activities were found to decrease more markedly in the freeze-dried plasmas than in the deep-frozen batches. Storage stability data at ambient temperature are not reported in this study [7]. The German Red Cross introduced a freeze-dried pathogen-inactivated SD plasma product in 1990. The product was examined to determine whether the quality was comparable to standard preparations. It was determined that freeze-dried SD plasma did not fulfill basic requirements. The time required to reconstitute the lyophilized product was too long. Also, the resultant pH values of the lyophilized/reconstituted plasma were close to the alkaline range; thus, considerable changes in blood gas and electrolyte levels were to be expected in the recipient [8]. In a separate study, the quality of three conventional fresh-frozen plasma preparations and one freeze-dried SD plasma preparation were compared [9]. Coagulation activity was significantly reduced in the freeze-dried SD plasma [9]. Storage stability data at ambient temperature are not reported in these studies [8] and [9]. In Thailand, freeze-dried plasma has been used by hemophilia patients as an in home treatment product since 1982. The chemical and coagulation properties of this product are reported nearly the same as FFP after reconstitution with sterile water [1]. The clinical effectiveness of this product has been shown in hemophilia patients with bleeding episodes. However, storage of the freeze-dried plasma product is still confined to 4° C. and requires refrigeration equipment [2].
Bakaltcheva et al. developed a method for production of freeze-dried whole plasma with increased stability at ambient temperature, physiological pH and osmolality [10]. Briefly, plasma supplemented with 60 mM sucrose, trehalose, mannitol, sorbitol or glycine was freeze-dried. The samples were subjected to forced degradation at 40 C for 10 days in order to quickly evaluate the effectiveness of the different stabilizers. Initial PT, APTT and TT values were 14.4 d 0.5 s, 31.4±1.5 s and 18.3±0.6 s, respectively. At the end of the degradation period, PT, APTT and TT were substantially prolonged. In the presence of glycine, at the end of the degradation period, PT, APTT and TT values remained close to the initial values and were 15.5±0.4 s, 35.7±0.9 s and 19.4±0.2 s, respectively. All tested stabilizers provided protection. Glycine, however, outperformed all tested polyols, providing superior preservation of plasma clotting properties. The process of freeze-drying caused a complete loss of plasma pCO2 (gas) and a substantial increase in plasma pH. Citric and ascorbic acid were found to be suitable pH adjusters for lyophilized/rehydrated plasma. However, the aim of this work was limited to the development of a method for production of freeze-dried whole plasma with increased stability, physiological pH and osmolality. A suitable pathogen inactivation method is to be combined with the lyophilization technology in order to increase the safety and availability of human plasma in rural areas and austere environments.
Gamma irradiation effectively inactivates all known blood-borne viruses and is currently explored vigorously as a pathogen inactivation method for plasma derivatives [11]. However, its application at virucidally effective doses (50 kGy) to frozen plasma products results in unacceptable losses in functional activity [11]. Ascorbate is the most commonly used protectant for plasma proteins against gamma irradiation [11-13]. However other antioxidants/stabilizers may be important when a high irradiation dose is applied. Grieb T et al. reported that the loss of protein activity after application of high dose gamma irradiation can be controlled [14]. Control is achieved by a combination of protection through the use of the antioxidant ascorbate and by freeze-drying to minimize the potential for generating free radicals.
Trauma induced coagulopathy is associated with an extremely high mortality. Transfusion of blood components remains the main treatment approach, with some recent studies advocating a more aggressive use of fresh frozen plasma (FFP) [15-16]. Most of the battlefield deaths take place before reaching a medical facility. Therefore, there is a clear need for the development of innovative and effective strategies for the early (pre-hospital) treatment of coagulopathy. One solution is to utilize freeze-dried, shelf-stable plasma, which has been subjected to a reliable pathogen inactivation method. However, such freeze-dried plasma has to be specifically designed to meet the needs of the trauma patient. Hemodilution, hypothermia, acidosis, tissue hypoperfusion and consumption of clotting factors are common reasons for the development of coagulopathy in trauma patients [17]. Once a stabilizer-supplemented freeze-dried plasma is liquefied upon reconstitution and administered to a patient, the stabilizer will be available to exercise its protective properties within the complex context of the specific injury or disease being treated. Therefore, when formulating freeze-dried plasma for use in the trauma care field, one should consider the complex state of a trauma patient and utilize stabilizers, which will support and not retard the recovery of the trauma patient.
There is a need to develop a method for production of freeze-dried whole plasma with increased stability at ambient temperature, physiological pH and osmolality.